A method for refining va palmitate

Abstract

This invention discloses a method for purifying VA palmitate. The purification method involves a two-step cooling and crystallization process to remove palmitic acid impurities from the reaction solution, followed by post-treatment to obtain purified VA palmitate. In the second cooling and crystallization step, nitrogen gas is bubbled through the bottom of the crystallizer, and the cooling rate and nitrogen flow rate are controlled to ensure a specific relationship between the cooling rate (ΔT') and the nitrogen flow rate (ΔV'). This invention solves the problems of low product yield and the risk of toxic solvent residues in existing technologies.

Description

Technical Field

[0001] This invention belongs to the field of separation and purification technology, and specifically relates to a purification method for VA palmitate. Background Technology

[0002] Vitamin A palmitate is a vitamin A ester derivative, considered to be the existing form of vitamin A compounds. It plays an important role in vision, the nervous system, and cell differentiation, and is an essential substance for normal human metabolism. Compared to other vitamin A derivatives, it is more stable under the influence of light, oxygen, and heat, and is widely used in food, pharmaceuticals, and nutritional supplements.

[0003] Currently, publicly available technologies primarily employ crystallization for the separation and purification of vitamin A palmitate. The existing technology, "Process Design for the Biocatalytic Production of Vitamin A Palmitate and its Industrial Production," discloses a crystallization-based separation method. This method first removes palmitic acid through cooling crystallization, but this process results in the loss of vitamin A palmitate. Secondly, acetone recrystallization is used for further separation and purification, but the overall product yield is only 86%. Furthermore, this method carries the risk of acetone residue, affecting the downstream applications of the product.

[0004] Current methods suffer from drawbacks such as operational complexity, low product yield, and the use of toxic reagents. A purification method for VA palmitate needs to be developed to address the problems of low product yield and the risk of toxic solvent residues inherent in existing technologies. Summary of the Invention

[0005] One of the objectives of this invention is to develop a purification method for VA palmitate, which efficiently removes impurities and improves product yield by controlling the coupling relationship between the cooling rate and the nitrogen gas introduction rate, thereby solving the problems of low product yield and risk of toxic solvent residue in the prior art.

[0006] To achieve the above-mentioned objectives, the technical solution of the present invention is as follows:

[0007] A method for purifying VA palmitate, wherein the purification method involves passing the reaction solution of VA palmitate to be treated through two steps of cooling and crystal growth to remove the impurity palmitic acid in the reaction solution, and then obtaining the purified VA palmitate through post-treatment.

[0008] In the second step of cooling and crystal growth, nitrogen gas is bubbled through the bottom of the crystallizer. The cooling rate and the nitrogen flow rate are controlled so that the cooling rate (ΔT') and the nitrogen flow rate (ΔV') satisfy the following relationship:

[0009] ΔV′=(-0.07ΔT′+2.2543)e 0.15*ΔT′

[0010] In the formula, ΔT' is 6~10℃ / h, and ΔV' is 4.5~7L / h, preferably 5.6~7L / h.

[0011] The inventors discovered in their research that the crystal size of palmitic acid, an impurity, affects the product yield during the refining process of VA palmitate products; specifically, larger palmitic acid crystals tend to coat the product during crystallization, leading to product loss. This invention effectively controls the cooling rate and nitrogen gas introduction rate to maintain a smaller palmitic acid crystal size, thereby reducing product loss and improving product yield.

[0012] Examples include, but are not limited to, the solutions listed in Table 1:

[0013] Table 1. Relationship between cooling rate and nitrogen injection rate

[0014] Cooling rate (ΔT') / (°C / h) Nitrogen gas flow rate (ΔV') / (L / h) Cooling rate (ΔT') / (°C / h) Nitrogen gas flow rate (ΔV') / (L / h) 6 4.5 8.5 5.9 6.5 4.8 9 6.3 7 5.0 9.5 6.6 7.5 5.3 10 7.0 8 5.6

[0015] In one embodiment of the present invention, the content of VA palmitate in the reaction solution is 15-45 wt%, preferably 25-45 wt%.

[0016] In one embodiment of the invention, the palmitic acid content in the reaction solution is 0.05-5 wt%, preferably 1-5 wt%.

[0017] In one embodiment of the present invention, the solvent of the reaction solution is an alkane organic solvent, preferably one or more of n-hexane, n-heptane, and n-octane.

[0018] In one embodiment of the present invention, the temperature of the reaction solution is 30-40°C, preferably 30-35°C.

[0019] In one embodiment of the present invention, the cooling rate of the first step of cooling and crystal growth is 15-20°C / h, cooling down to 0-5°C, preferably cooling down to 0-3°C, and the crystal growth time is 2-4h, preferably 2-3h.

[0020] In one embodiment of the present invention, the cooling rate of the second-stage cooling and crystal growth is 6-10°C / h, the temperature is cooled to -5-10°C, preferably to -8-10°C, and the crystal growth time is 2-4h, preferably 2-3h.

[0021] In one embodiment of the present invention, the post-treatment includes filtration, washing, and solvent removal; preferably, the washing temperature is -5 to -10°C, more preferably -8 to -10°C, and the mass ratio of washing solvent to filtrate is 1-3:1, more preferably 2-3:1; preferably, the solvent removal temperature is 20 to 40°C, more preferably 30 to 40°C, and the pressure is 1 to 5 kPaA, more preferably 1 to 3 kPaA.

[0022] Another object of the present invention is to provide a VA palmitate.

[0023] A VA palmitate, wherein the VA palmitate is prepared by the above-described purification method, and the VA palmitate contains palmitic acid at a mass content of 50-2000 ppm.

[0024] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0025] 1) Effective control of the coupling between cooling rate and nitrogen gas introduction rate resulted in a VA palmitate yield of over 99%, higher than the 86% separation yield in existing methods; meanwhile, the palmitic acid content in the obtained product was 50-2000 ppm, meeting the product specification requirements.

[0026] 2) Low-toxicity organic solvents are used in the refining process to reasonably avoid the risk of acetone residue in existing technical solutions. Detailed Implementation

[0027] To facilitate understanding of the present invention, the following description, in conjunction with embodiments, will further illustrate the invention. It should be understood that the following embodiments are merely for a better understanding of the invention and do not imply that the invention is limited to these embodiments.

[0028] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items. Locational terms such as top and bottom mentioned or that may be mentioned in this specification are defined relative to the illustrated construction and are relative concepts, and therefore may vary depending on their location and usage.

[0029] Main raw material sources

[0030] The reagents used in this invention, namely n-hexane, n-heptane, n-octane, ethanol, and methanol, were all purchased from Beijing Innocare Technology Co., Ltd., and were of analytical grade.

[0031] Palmitic acid was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd.

[0032] VA palmitate was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd.

[0033] Main testing methods

[0034] The analytical methods used for the content of VA palmitate and palmitic acid in this invention are as follows: High performance liquid chromatography (HPLC) using an Agilent LC-1200 column (4.6 × 250 mm, 5 μm). VA acetate, VA palmitate, and VA alcohol were detected using a UV detector at a wavelength of 326 nm, with quantification using the external standard method. Palmitic acid was quantified using an ELSD detector with the external standard method. The corresponding chromatographic detection conditions are shown in Table 1.

[0035] Table 2 Chromatographic Detection Conditions

[0036]

[0037] Product efficacy testing was performed according to the method described in GB 29943-2013, using an ultraviolet spectrophotometer (Agilent Cary 3500) to detect the absorbance value of the sample at 326 nm, and calculating it based on the empirical formula described in the standard.

[0038] Prepare solutions containing different concentrations of palmitic acid and VA palmitate:

[0039] Solution 1: Using a 250mL three-hole glass reaction flask, after purging with nitrogen, weigh 15g VA palmitate, 0.05g palmitic acid and 84.95g n-heptane, stir and mix evenly, and set aside.

[0040] Solution 2: Using a 250mL three-hole glass reaction flask, after purging with nitrogen, weigh 25g VA palmitate, 1g palmitic acid and 74g n-hexane, stir and mix evenly, and set aside.

[0041] Solution 3: Using a 250mL three-hole glass reaction flask, after purging with nitrogen, weigh 45g VA palmitate, 5g palmitic acid and 50g n-octane, stir and mix evenly, and set aside.

[0042] Solution 4: The VA palmitate esterase catalytic reaction solution was prepared according to the method described in patent CN116023314A. The VA palmitate content in the reaction solution was 32% / wt, the palmitic acid content was 1.8% / wt, and the solvent was n-hexane.

[0043] Example 1

[0044] Prepare the product by refining solution 1.

[0045] Take 50g of VA palmitate solution 1 and keep it at 30℃. In the first stage, the stirring rate is 100rpm and the cooling rate is 15℃ / h, cooling to 0℃ for 3 hours. In the second stage, the stirring rate is 100rpm and the cooling rate is 6℃ / h, cooling to -5℃ for 2 hours. During the second stage cooling and crystal growth process, nitrogen gas is introduced into the bottom of the crystallizer at a cooling rate of 6℃ / h. The calculated value on the right side of the equation is 4.5, therefore the nitrogen gas flow rate is 4.5L / h. A glass-jacketed funnel with 800-mesh filter cloth (19μm pore size) at the bottom is used. After filtration at 50KPa(A), an equal mass of ethanol (at -5℃) is added to wash the filter cake, maintaining the temperature at -5℃ throughout the washing process. After phase separation, the upper alkane phase is taken for desolventizing at 20℃ and 1KPa(A). 7.45 g of VA palmitate product was obtained, with a product yield of 99.3%. The potency of the VA palmitate product was determined to be 1.8 million IU / g, and the residual palmitic acid content was 151 ppm.

[0046] Example 2

[0047] The product is prepared by refining solution 2.

[0048] Take 50g of VA palmitate solution 2 and keep it at 35℃. In the first stage, the stirring rate is 100rpm and the cooling rate is 17℃ / h, cooling to 3℃ for 2 hours. In the second stage, the stirring rate is 100rpm and the cooling rate is 8℃ / h, cooling to -8℃ for 3 hours. During the second stage cooling and crystal growth process, nitrogen gas is introduced into the bottom of the crystallizer at a cooling rate of 8℃ / h. The calculated value on the right side of the equation is 5.6, indicating a nitrogen gas flow rate of 5.6L / h. A glass-jacketed funnel with 800-mesh filter cloth (19μm pore size) at the bottom is used for filtration at 50KPa(A). After filtration, ethanol (3 times the mass of the filtrate, -8℃) is added to wash the filter cake, maintaining the temperature at -8℃ throughout the washing process. After phase separation, the upper alkane phase is used for desolventizing at 30℃ and 3KPa(A). 12.45 g of VA palmitate product was obtained, with a product yield of 99.6%. The potency of the VA palmitate product was determined to be 1.83 million IU / g, and the residual palmitic acid content was 56 ppm.

[0049] Example 3

[0050] The product is prepared by refining the solution 3.

[0051] Take 50g of VA palmitate solution 3 and keep it at 40℃. In the first stage, the stirring rate is 100rpm and the cooling rate is 20℃ / h, cooling to 5℃ for 4h. In the second stage, the stirring rate is 100rpm and the cooling rate is 10℃ / h, cooling to -10℃ for 4h. During the second stage cooling and crystal growth process, nitrogen gas is introduced into the bottom of the crystallizer at a cooling rate of 8℃ / h. The calculated value on the right side of the equation is 7, therefore the nitrogen gas flow rate is 7L / h. A glass-jacketed funnel with 800-mesh filter cloth (19μm pore size) at the bottom is used for filtration at 50KPa(A). After filtration, methanol (at -10℃) with twice the mass of the filtrate is added to wash the filter cake, maintaining the temperature at -10℃ throughout the washing process. After phase separation, the upper alkane phase is taken for desolventizing at 40℃ and 5KPa(A). 22.37 g of VA palmitate product was obtained, with a product yield of 99.4%. The potency of the VA palmitate product was determined to be 1.81 million IU / g, and the residual palmitic acid content was 128 ppm.

[0052] Example 4

[0053] The product is prepared by refining the solution 4.

[0054] Take 50g of VA palmitate solution 4 and prepare the product according to the process described in Example 1 to obtain 15.89g of VA palmitate product, with a product yield of 99.3%. The potency of the VA palmitate product was determined to be 1.81 million IU / g, and the residual palmitic acid content was 98ppm.

[0055] Comparative Example 1

[0056] Compared with Example 3, the only difference is that nitrogen gas is not introduced for bubbling.

[0057] Take 50g of VA palmitate solution 3 and prepare the product according to the process described in Example 3, except that nitrogen gas was not introduced. 19.26g of VA palmitate product was obtained, with a product yield of 85.6%. The potency of the VA palmitate product was determined to be 1.73 million IU / g, and the residual palmitic acid content was 2300 ppm.

[0058] Comparative Example 2

[0059] The cooling rate is not within the preferred range.

[0060] Take 50g of VA palmitate solution 2 and prepare the product according to the process described in Example 2, the difference being that the cooling rates in the first and second stages are 64℃ / h and 22℃ / h, respectively (not within the preferred range); the nitrogen gas introduction rate is still 5.6L / h. 10.01g of VA palmitate product was obtained, with a product yield of 80.1%. The potency of the VA palmitate product was determined to be 1.71 million IU / g, and the residual palmitic acid content was 2900 ppm.

[0061] Comparative Example 3

[0062] Compared with Example 2, the only difference is that the cooling rate in the second stage does not satisfy the relationship between the nitrogen gas flow rate and the nitrogen gas flow rate.

[0063] Take 50g of VA palmitate solution 1 and prepare the product according to the process described in Example 1. The difference is that the cooling process in the second stage is not related to the nitrogen gas flow rate. At this time, the cooling rate is 6℃ / h, and the calculated value on the right side of the equation is 4.5, while the nitrogen gas flow rate is 6L / h. 6.78g of VA palmitate product was obtained, with a product yield of 90.3%. The potency of the VA palmitate product was determined to be 1.76 million IU / g, and the residual palmitic acid was 2100ppm.

[0064] It is readily understood that the above embodiments are merely illustrative examples for clear explanation and do not imply that the invention is limited thereto. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A method for purifying VA palmitate, characterized in that, The purification method involves passing the reaction solution of VA palmitate to be treated through two steps of cooling and crystal growth to remove the impurity palmitic acid in the reaction solution, and then obtaining purified VA palmitate through post-treatment. In the second step of cooling and crystal growth, nitrogen gas is bubbled through the bottom of the crystallizer. The cooling rate and the nitrogen flow rate are controlled so that the cooling rate (ΔT') and the nitrogen flow rate (ΔV') satisfy the following relationship: AV' = (-0.07 ΔT' + 2.2543) e 0.15*ΔT′ In the formula, ΔT' is 6~10℃ / h, and ΔV' is 4.5~7L / h, preferably 5.6~7L / h.

2. The refining method according to claim 1, characterized in that, The content of VA palmitate in the reaction solution is 15-45 wt%, preferably 25-45 wt%; And / or, the palmitic acid content in the reaction solution is 0.05-5 wt%, preferably 1-5 wt%; And / or, the solvent of the reaction solution is an alkane organic solvent, preferably one or more of n-hexane, n-heptane, and n-octane; And / or, the temperature of the reaction solution is 30–40°C, preferably 30–35°C.

3. The refining method according to claim 1 or 2, characterized in that, The cooling rate for the first step of cooling and crystal growth is 15-20℃ / h, cooling down to 0-5℃, preferably to 0-3℃, and the crystal growth time is 2-4h, preferably 2-3h. And / or, the cooling rate of the second-stage cooling and crystal growth is 6 to 10 °C / h, cooling to -5 to -10 °C, preferably cooling to -8 to -10 °C, and the crystal growth time is 2 to 4 h, preferably 2 to 3 h.

4. The refining method according to any one of claims 1-3, characterized in that, The post-processing includes filtration, washing, and solvent removal; Preferably, the washing temperature is -5 to -10°C, more preferably -8 to -10°C, and the mass ratio of washing solvent to filtrate is 1-3:1, more preferably 2-3:1; Preferably, the desolventizing temperature is 20–40°C, more preferably 30–40°C, and the pressure is 1–5 kPaA, more preferably 1–3 kPaA.

5. A VA palmitate prepared by the purification method according to any one of claims 1-4, wherein the VA palmitate contains palmitic acid in a mass content of 50-2000 ppm, preferably 50-160 ppm.

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